Eicosapentaenoic acid (hereinafter referred to as “EPA”) and docosahexaenoic acid (hereinafter referred to as “DHA”) are the polyunsaturated fatty acids known in particular for their numerous physiological functions, including preventive effects against adult diseases such as arteriosclerosis and thrombosis, anticancer effects and learning reinforcement effects, and they are often utilized in drugs and special healthy foods and/or supplements. Recently, however, the physiological functions of other polyunsaturated fatty acids have been the subject of increasing attention.
Arachidonic acid is one of these polyunsaturated fatty acids and constitutes approximately 10% of the fatty acids composing important organs such as the blood and liver (for example, the compositional ratio of fatty acids in human blood phospholipids is 11% arachidonic acid, 1% eicosapentaenoic acid and 3% docosahexaenoic acid). As a major constituent of cell membranes it is involved in regulating membrane fluidity, and performs various roles in biometabolism while also serving as an important direct precursor to prostaglandins. A recent area of attention has been the role of arachidonic acid as an infant nutrient and its presence as a constituent fatty acid of endogenous cannabinoids (2-arachidonoyl monoglycerol, anandamide) which exhibit a neurostimulating action. Consumption of linoleic acid-rich foods usually results in conversion to arachidonic acid, but direct ingestion of arachidonic acid in the form of triglycerides is preferred because of a reduced function of the enzymes involved in biosynthesis in adult disease patients and those at risk, infants and the elderly who, as a result, tend to be deficient in arachidonic acid.
Fish oil is an abundant source of EPA and DHA, but dihomo-γ-linolenic acid, arachidonic acid and 4,7,10,13,16-docosapentaenoic acid (22:5 ω6) are almost unobtainable from conventional oil and fat sources. At the current time, it is common to use oils/fats or triglycerides whose constituent fatty acids are polyunsaturated fatty acids and which are obtained by fermentation with microbes. For example, according to one proposed method, various microbes capable of producing oils/fats or triglycerides containing arachidonic acid as the constituent fatty acid are cultured to yield the oils/fats or triglycerides containing arachidonic acid as the constituent fatty acid. Such methods include those for obtaining arachidonic acid-rich oils/fats or triglycerides using fungi of the genus Mortierella (see, for example, Japanese Unexamined Patent Publication No. 63-44891 and Japanese Unexamined Patent Publication No. 63-12290). Triglycerides containing fermentation-produced arachidonic acid as the constituent fatty acid are used for purposes requiring arachidonic acid, such as in the field of infant nutrition and, particularly, in modified milk.
Methods of adding arachidonic acid-containing oils or fats to modified milk have been disclosed in the field of infant nutrition (see Japanese Unexamined Patent Publication No. 11-151075, Japanese Unexamined Patent Publication No. 10-191886). The arachidonic acid-containing oils or fats used as additives are produced by fungi, and from a molecular standpoint are characterized by comprising 6-24% AAA (a triglyceride with 3 residues of arachidonic acid in the molecule). A higher arachidonic acid content is known to result in a higher AAA concentration (see, for example, Jim-Wen Liu et al., In vitro hydrolysis of fungal oils: distribution of arachidonic acid-containing triacylglycerol molecular species, J. Am. Oil Chem. Soc., 75, pp. 507-510(1998)).
Oils and fats containing a high concentration of AAA differ from vegetable oils and fats in being resistant to the action of human digestive enzymes (pancreatic lipases) under physiological conditions and, therefore, such oils and fats are not readily digested and absorbed by infants or elderly with low pancreatic lipase activity (see, for example, Jim-Wen Liu et al.).
As the arachidonic acid content of human breast milk is 0.5% of the total fatty acids (see, for example, Christie, W. W., “The positional distribution of fatty acids in triglycerides” in Analysis of oils and fats, Edited by Hamilton, R. J. and Russell, J. B., pp. 313-339, Elsevier Applied Science, London (1986)), presumably there is a higher probability of one arachidonic acid residue per triglyceride molecule rather than greater arachidonic acid condensation per molecule as in AAA described above. Consequently, addition of arachidonic acid-containing oils and fats obtained by fermentation with fungi to modified milk simply on the basis of the fatty acid content is not desirable.
Many attempts have been made to enzymatically alter oils and fats to enhance their properties (solubility in the mouth, crystallinity, heat resistance). Most have involved enzymatic modification of vegetable oils or fats, such as enzymatic synthesis of cacao substitute oils (see, for example, Japanese Unexamined Patent Publication No. 55-71797, Japanese Unexamined Patent Publication No. 58-42697). The production technology disclosed here is for vegetable oils and fats that exhibit high lipase reactivity, whereas oils and fats rich in polyunsaturated fatty acids with poor reactivity are poorly suitable in terms of lipase reactivity.
Oils/fats or triglycerides rich in polyunsaturated fatty acids have not been used for enzymatic modification because of their poor enzyme reactivity, but it has been attempted to produce lipids with polyunsaturated fatty acids structure using medium-chain fatty acid- and polyunsaturated fatty acids-rich oils and fats, by using altered immobilized enzymes (see Japanese Unexamined Patent Publication No. 8-214891, PCT/JP02-06702). According to this method, the fatty acid at the 1- and/or 3-positions of the triglyceride are replaced with octanoic acid, thereby releasing the free polyunsaturated fatty acids. Polyunsaturated fatty acids-rich oils or fats are therefore not expected products. Another disadvantage is that further purification techniques such as precision distillation and the like are required to remove the free fatty acids.
Fujimoto et al. have studied in detail the effects of oil/fat structure on the oxidative stability of polyunsaturated fatty acids, in light of the fact that fish oils (particularly oils rich in EPA and DHA) readily oxidize and adequate oxidative stability cannot be achieved with addition of antioxidants (see, for example, Fujimoto, K., “Effects of oil and fat structure on oxidation stability of polyunsaturated fatty acids”, Science and Industry, 75, pp. 53-60, Osaka Industrial Research Association (2001)). An improvement in oxidative stability was discovered by adding special fats (C8 and C14 triglycerides of medium chain fatty acids) to sardine oil (rich in polyunsaturated fatty acids) and conducting transesterification with position non-specific type lipases, and upon examining the ease of oxidation of the chemically synthesized polyunsaturated fatty acids triglyceride EEE (tri-EPA), and comparing types with the polyunsaturated fatty acids dispersed among the molecules and with a higher condensation in the same molecule, the dispersed type was confirmed to be satisfactory.
Transesterification has been disclosed as a method of stabilizing oils and fats in order to prevent oxidation of polyunsaturated fatty acids in fish oils and the like (see Japanese Unexamined Patent Publication No. 6-287593). However, the polyunsaturated fatty acids in fish oils and the like have low reactivity for the lipases used in transesterification reactions. According to this method, therefore, vegetable oils or fats with high lipase reactivity are used in a large amount to dilute one or more polyunsaturated fatty acids-containing oils purified from fish oil or the like in order to ensure lipase reactivity. As a result, it is not possible to achieve an increased content of polyunsaturated fatty acids in the stabilized oils and fats.